Probing the sulfhydryl groups of nuclear matrix proteins with 6-iodoacetamidofluorescein.

Abstract

Rat liver nuclear matrices were reacted with the fluorescent dye 6-iodoacetamidofluorescein and the matrix proteins were then separated by one and two-dimensional polyacrylamide gel electrophoresis. Upon transillumination with U.V. light it was possible to see that several proteins had reacted with the dy, thus indicating the presence of free -SH groups. This labelling technique allowed the detection of a large number of proteins, being several folds more sensitive than conventional Coomassie Blue staining, as demonstrated by two-dimensional electrophoretical separation. If nuclear matrices were treated with reducing agents before being reacted with 6-iodoacetamidofluorescein, the fluorescence increased with about the same intensity in all the protein bands. It is proposed that 6-iodoacetamidofluorescein can be used as a specific and very sensitive probe to study the -SH groups of nuclear matrix proteins.